Diesel fuel is any fuel used in diesel engines and includes both petrodiesel and biodiesel. Petrodiesel is a specific fractional distillate of fossil fuel oil. It is comprised of about 75% saturated hydrocarbons and 25% aromatic hydrocarbons. Biodiesel is not derived from petroleum but from vegetable oil or animal fats and contains long chain alkyl esters. Biodiesel is made by the transesterification of lipids (e.g., spent vegetable oil from fryers or seed oils) with an alcohol and burns cleaner than petrodiesel. Biodiesel can be used alone or mixed with petrodiesel in any amount for use in modern engines.
Kerosene is a combustible hydrocarbon that is also a specific fractional distillate of fossil fuel and contains hydrocarbons having 6 to 16 carbon atoms. Kerosene has a heat of combustion comparable to that of petrodiesel and is widely used in jet fuel to power jet engines and for heating in certain countries. Kerosene-based fuels can also be burned with liquid oxygen and used as rocket fuel (e.g., RP-1).
Reliance on petroleum-derived fuels has depleted the supply of natural resources and has required increased reliance on imported gasoline and diesel products. In addition, the burning of petroleum-based fuels has increased the amount of greenhouse gasses (e.g., carbon dioxide and methane) in the atmosphere that is contributing to the gradual warming of the earth's climate.
Fuels, such as biodiesel, that are made from animal or vegetable products burn cleaner than petroleum-derived fuels and do not produce a net increase in greenhouse gases. Furthermore, they are a sustainable energy source and have the potential to reduce the United States' reliance on imported petroleum-based products. However, there is a concern that using land to produce fuel crops rather than food crops will contribute to world hunger.
Fatty acids are the principle component of cell membranes and are used by nearly all organisms as a primary source of energy storage. Fatty alcohols are the reduction products of fatty acids and, like fatty acids, can be produced enzymatically by cultured cells. Fatty alcohols can be reacted with acids to form ester compositions similar to those present in biodiesel fuel, or reduced to form kerosene-like compositions, or hydrocarbon compositions similar to petrodiesel. Enzymes that convert fatty acyl-thioester substrates (e.g., fatty acyl-CoA or fatty acyl-ACP) to fatty alcohols are commonly referred to as fatty alcohol forming acyl-CoA reductases or fatty acyl reductases (“FARs”).
PCT Publication No. WO 2007/136762 discloses genetically engineered microorganisms for the production of fatty acid derivatives and methods of their use.
PCT Publication No. WO 2009/140695 discloses compositions comprising cyanobacterial genes encoding enzymes involved in hydrocarbon biosynthesis and methods of using then in the production of aldehydes and alcohols.
Steen et al., 2010, Nature 463:559-563 discloses the engineering of E. coli to produce specific fatty esters, fatty alcohols and waxes directly from simple sugars.
Schirmer, 2009, Current Opinion in Microbiology 12:274-281 describes the fermentative and nonfermentative metabolism of heterotrophic microorganisms used in the production of fuel-like molecules, biocatalysts that convert metabolic intermediates into fuel-like molecules, and the parameters that govern the cost effective production of such fuel-like molecules.
U.S. Pat. No. 5,370,996 and Metz et al. (2000)Plant Physiology 122:635-644 disclose isolation and characterization of a fatty acyl reductase (FAR) enzyme from the desert shrub Simmondsia chinensis, more commonly known as jojoba.
Moto et al. (2003) Proc. Nat'l Acad. Sci. USA 100(16):9156-9161 discloses the isolation and characterization of a FAR enzyme from the silk moth Bombyx mori. 
Reiser et al. (1997) J. Bacteriol. 179(9):2969-2975 discloses the isolation and characterization of a fatty acyl CoA reductase enzyme from the wax ester producing bacterium Acinetobacter calcoaceticus that reduces a fatty acyl-CoA substrate with chain lengths from C14 to C22 to the corresponding fatty aldehyde, requiring a dehydrogenase enzyme for conversion of the fatty aldehyde to the fatty alcohol.
In theory, these FAR enzymes could be expressed in heterologous hosts as a means of producing a non-petroleum-based, renewable source of fatty alcohols or derivative compositions for use in biofuels. However, when expressed in heterologous hosts such as E. coli and yeast, the yields of fatty alcohols obtained have been insufficient for certain applications. In addition, at most, only a small fraction of the produced fatty alcohols are secreted by the microorganisms, increasing substantially the cost of purification.
Accordingly, there remains a need in the art for enzymes such as FAR enzymes that can be used to efficiently produce fatty alcohols for use in industrial applications such as, but not limited to applications in the food industry, cosmetic industry, medical industry, and fuels industry.